We have undertaken time-resolved Forster resonance energy transfer (FRET) and molecular dynamics simulations to analyze conformations and conformational heterogeneity of an analogue of leucine enkephalin in solution and in the presence of sodium dodecyl sulfate (SDS) micelles. Enkephalins are opioid pentapeptides that interact with opioid receptors in the central nervous system. We used time-correlated single-photon counting to detect energy transfer between the N-terminal tyrosine and a tryptophan residue substituted for phenylalanine at the 4 position. FRET from Tyr to Trp was measured over a temperature range from 5 to 55 degrees C in aqueous solution. By taking into account Tyr rotamer interconversion rates measured previously, we determined average distances between Tyr and Trp for the two populated rotameric conformations of Tyr. Molecular dynamics simulations (100 ns) support this analysis and indicate extensive conformational heterogeneity. The simulations also predict that the FRET orientational factor is correlated with the Tyr-Trp separation. Failure to account for the correlation between orientation and distance results in errors that appear to be largely offset in the leucine enkephalin analogue (YGGWL) by a weighting bias inherent in the R-6 dependence of the energy-transfer rate. The Tyr lifetimes decrease upon titration of the peptides with SDS, indicating formation of compact conformations of the peptide in the micelle environment. This result is consistent with the conjecture that the lipid environment may induce formation of bioactive conformations of the peptide.